Paper point of care

A team of U.S. researchers has
developed a postage stamp-sized diagnostic for hepatotoxicity and has used it
to measure liver enzyme levels in human blood samples.1Diagnostics for All has exclusively licensed the device and is running
field tests in Vietnam.

Blood tests for monitoring liver status in at-risk
individuals receiving tuberculosis (TB) or HIV therapies are a standard
practice.2-4 In the developing world, however, logistical
limitations can make it difficult and costly to monitor therapy-associated
hepatotoxicity.

As a result, there is a need for liver toxicity tests
that can be rapidly implemented and interpreted in a resource-limited setting
and that are affordable, stable and easy to use.5

Prior studies have shown that paper-based microfluidic
devices are useful for conducting basic enzymatic and immunoassay tests.6-9
Such devices require no external pumps, instrumentation or power and are
portable and disposable, making them suitable for use in the developing world.
However, the devices have yet to be validated with actual patient samples.

Thus, a team of researchers led by Nira Pollock, Jason
Rolland and George Whitesides set out to design a paper-based device that
measured liver enzyme levels in blood from patients receiving HIV and/or TB
therapy and that required only the unaided eye to detect and interpret results.
To do that, they relied on basic microfluidics technology previously developed
in Whitesides' laboratory.10, 11

The study was performed on U.S. patients by U.S.
clinicians and researchers.

When a drop of whole blood was applied to an opening in
the cover of the device, blood cells were retained in the separation membrane
while plasma diffused into the other layers and reacted with reagents. That
reaction generated a color readout that indicated the presence of aspartate
aminotransferase (AST) and alanine
aminotransferase (ALT) in the plasma. AST
and ALT are two enzymes commonly associated with liver injury.

The intensity of the color readout was optimized to fall
within one of three clinically relevant ranges: <3´,
3´-5´ and >5´
normal AST and ALT levels.

The surface of the paper also included three control
zones to monitor device performance. The entire test was completed in about 15
minutes.

The researchers next tested their device on 233 blood
samples from patients with HIV and TB who had a range of AST and ALT
concentrations. The device was >90%
accurate using both serum and whole-blood samples.

The authors wrote that their method "can ultimately
be produced at a very low cost-on the order of <$0.10 per test." Current electronic gold standard
tests cost $4.

Results were published in Science Translational
Medicine.

"The paper provides the first published example of
clinical tests using paper-based microfluidic devices and provides convincing
evidence that the basic premise of paper microfluidics is sound and worth
substantial development," said Scott Phillips, assistant professor of
chemistry at Pennsylvania State University.

Earlier this year, Phillips, Whitesides and colleagues
published in Analytical Chemistry data on a paper-based microfluidic
device that measured alkaline phosphatase, AST and total
protein levels in nonclinical samples.12

In contrast to much previous work in paper-based
diagnostics and assays, the authors "considered logistical constraints
from the outset in designing their test, including cost factors, device storage
and potential user error," said Barry Lutz.

They also "chose to align their test with current
clinical practice, which can greatly increase the likelihood of adoption,"
he added.

Lutz and colleagues Paul Yager and Elain Fu at the University of Washington
are developing paper devices for amplified immunoassays and nucleic acid
amplification tests,13 with the goal of creating methods that "perform
sophisticated biochemical tests in a format that is as easy to use as a
pregnancy test."

Lutz and Fu are research assistant professors of
bioengineering at the University of Washington. Yager is professor and chair of
bioengineering at the university.

Good morning, Vietnam

Kenneth Hawkins, program officer and
research scientist in the diagnostics group at PATH
(Program for Appropriate Technology in Health), said the next step should be "a
field-based study in one of the target markets in the developing world. The
results could be very different with a different patient population and when
performed by a representative group of operators in the target market."

The color reading of the device "is subjective, and
the device's true ability to quantify results must be proven," added
Robert Jenison, CTO at Great Basin Corp. "A
reader device would help tremendously."

Great Basin is developing a battery-powered instrument
for use in the developing world to detect mutations in TB bacteria.

"It'll be interesting to test the outcome of the
assay if each patient is asked to read his or her own result," said
Phillips. "The ideal point-of-care device would be one that provides a
readout that is sufficiently unambiguous that anyone can read and interpret it."

Earlier this year, Diagnostics for All and Beth Israel
partnered with PATH and The Hospital for Tropical Diseases
(HTD) in Vietnam to carry out a preliminary study of the paper-based
diagnostic, co-corresponding author Pollock told SciBX. "We are
evaluating the performance of the diagnostic in finger-stick testing of 600
patients from the HTD HIV clinic," she said.

Initial data are expected in the next few months. The
field study results "will provide data on how well local clinicians are
able to read and interpret the test. Initial adoption of the test would likely
be in patient clinics where the test would be administered by a trained
healthcare provider," Pollock added.

"Additional next steps in the work include
long-term stability studies and development of quality manufacturing systems at
Diagnostics for All for large-scale production of validated lots of the device,"
said co-corresponding author Rolland.

The researchers also are interested in using companion
reader devices, said Rolland. "A particularly attractive means of doing
this is to use a smart phone able to recognize and interpret colorimetric
tests. This capability is something we plan to pursue in the next 1-2 years,"
he said.

The findings are covered by patent applications that are
exclusively licensed to Diagnostics for All from Harvard.

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